Atomic force microscope (AFM) combined with the ultramicrotome: a novel device for the serial section tomography and AFM/TEM complementary structural analysis of biological and polymer samples
Atomic force microscope (AFM) combined with the ultramicrotome: a novel device for the serial section tomography and AFM/TEM complementary structural analysis of biological and polymer samples
复制标题
DOI：
10.1111/j.1365-2818.2007.01773.x
复制
发表时间：
2007-06-01
影响因子：
2
通讯作者：
Matsko, Nadezda B.
中科院分区：
文献类型：
--
作者：
Efimov, Anton E.;Tonevitsky, Alexander G.;Matsko, Nadezda B.
介绍了一种基于扫描探针显微镜（SPM）（NT-MDT NTEGRA SPM）和超薄切片机（Leica UC 6 NT）集成的新设备（NTEGRA Tomo）。这种集成使得能够在超薄切片机切片程序的每个切片循环之后立即直接监测块面表面。因此，该设备可以应用于广泛的生物和聚合物材料的连续切片断层摄影。切片/扫描周期的自动化允许每小时获取多达10个连续切片层图像。它还允许在一个测量会话内从几十个层图像重建3D纳米断层摄影图像。层的厚度可以从20到2000 nm变化，并且可以通过其在水中的干涉颜色直接控制。此外，具有纳米分辨率的NTEGRA Tomo是缩小和突出样品体积内特别感兴趣区域的有效仪器。对于包埋的生物样品，SPM的最终分辨率主要取决于样品制备过程中生物材料大分子的保存质量。对于大多数聚合物材料，它可以与透射电子显微镜（TEM）相媲美。NTEGRA Tomo可以常规地收集AFM和TEM图像。生物或聚合物样品的块面通过AFM研究，而最后的切片在染色程序后用TEM分析。使用这两种超微结构方法的组合，用于分析相同的特定细胞器或聚合物成分，导致AFM/TEM图像解释的突破。最后，可以揭示物体超微结构的新的互补方面。
A new device (NTEGRA Tomo) that is based on the integration of the scanning probe microscope (SPM) (NT-MDT NTEGRA SPM) and the Ultramicrotome (Leica UC6NT) is presented. This integration enables the direct monitoring of a block face surface immediately following each sectioning cycle of ultramicrotome sectioning procedure. Consequently, this device can be applied for a serial section tomography of the wide range of biological and polymer materials. The automation of the sectioning/scanning cycle allows one to acquire up to 10 consecutive sectioned layer images per hour. It also permits to build a 3-D nanotomography image reconstructed from several tens of layer images within one measurement session. The thickness of the layers can be varied from 20 to 2000 nm, and can be controlled directly by its interference colour in water.Additionally, the NTEGRA Tomo with its nanometer resolution is a valid instrument narrowing and highlighting an area of special interest within volume of the sample. For embedded biological objects the ultimate resolution of SPM mostly depends on the quality of macromolecular preservation of the biomaterial during sample preparation procedure. For most polymer materials it is comparable to transmission electron microscopy (TEM).The NTEGRA Tomo can routinely collect complementary AFM and TEM images. The block face of biological or polymer sample is investigated by AFM, whereas the last ultrathin section is analyzed with TEM after a staining procedure. Using the combination of both of these ultrastructural methods for the analysis of the same particular organelle or polymer constituent leads to a breakthrough in AFM/TEM image interpretation. Finally, new complementary aspects of the object's ultrastructure can be revealed.